LOCUS       sequence-379183-        9437 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses NG-tagged ANME2cE20 MCR in Methanococcus maripaludis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9437)
  AUTHORS   Shao N, Fan Y, Chou CW, Yavari S, Williams RV, Amster IJ, Brown SM, 
            Drake IJ, Duin EC, Whitman WB, Liu Y
  TITLE     Expression of divergent methyl/alkyl coenzyme M reductases from 
            uncultured archaea.
  JOURNAL   Commun Biol. 2022 Oct 20;5(1):1113. doi: 10.1038/s42003-022-04057-6.
  PUBMED    36266535
REFERENCE   2  (bases 1 to 9437)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Commun 
            Biol."; date: "2022-10-20"; volume: "5(1):1113. doi"; pages: " 
            10.1038/s42003-022-04057-6"
COMMENT     Sequence Label: Ppst93-ANME2cE20 MCR-NG
FEATURES             Location/Qualifiers
     source          1..9437
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2922..2945
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(2955..2978,2979..3014,3015..3038)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     primer_bind     complement(6882..6901)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7001..7023
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7061..7079)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7147..7251
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7252..7320,7321..8112)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7470..7489)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8283..8871
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8772..8791
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttttttatat attatattat atttaacctt gtattgatta ttgaataata tttattatta
       61 ctttttttga tttcttgtaa aaaatcgaac atattttaca tattgtaaaa aataaaaaag
      121 atttaaaaaa agtgtccgaa aaagttaaac aaacattatt attaaaatgt tagtttaata
      181 tgtatcttga tatgctcttc tttctttcat gcttaaactt gtataactgt atgctttttt
      241 catttccttt attttgtcct ttcctttctt tgttatctgg taatatcctt ccttctttcc
      301 tgcatacttc aaaaatccaa ggttttttaa gttatcgaga taagaattac tagatgaatt
      361 cgcgcccggt tctagattta ttcataaaat aggaactaaa ataccaaaat agaaacattt
      421 atatacattt gatgtgtagg gtataccata aacctgggag gtgtctcata tggcagatac
      481 aattgatctt tacgatgata gaggtaaaaa acttaaagga gatgttgatt tacaagcagt
      541 ttcaccatta aaaaactcag caattcttgg aatggttaac acagttaaaa gaactgttgc
      601 agttaacctt gcaggaattg aaaaagcatg taaaaacgca tcatacggag gacaatcaag
      661 aaatattcca ggaagagaag ttgatattga tccaactgca aaagcagata aaattgcagc
      721 aagagttaaa gaacttattc aggttgaaaa aggagatgat acagaagtta ctgttttagg
      781 tggaggaaaa tttcttagag ttgcagcacc atcaagaaga attgaagcag gagcagaata
      841 tgttgcagga atgacatgta ctgcagcagc attaacagaa gcacttagag aagaatacgg
      901 acttggaatg tatgatactc catacgttaa aaatgcagtt tggggaacat acccacaaac
      961 tatggatatg aaaggtggaa acgttttatc agttctttca attccacaga atgatgaagg
     1021 acttggattt gcacttagaa acattatggc aaaccacctt gcaatgcttt cacaaagaaa
     1081 cgcaatgaac tgtgcagcaa tttcatcaat tttagaacac tgtggagttt ttgaaatggg
     1141 acaggcaatt ggactttttg aaagatacca acttttagca ttagcatacc agggacttaa
     1201 cgcaaacaac atggtttacg aaatgacaaa aaataatgga aaaacaggaa ctattggaac
     1261 agttgttcaa gaaactgttg aaagagcaat tgatgatgga gttattgcag ttgataaaac
     1321 tatgccatca ggatacaaag tttacaaagc aaacgatgtt tcaatgtgga atgcatattg
     1381 tgcagcagga acaatggcat caactatggt taattgtgga gcattaagag gagcacaggc
     1441 agtttcatca acacttcttt actttaacga tatgattgaa aaagaaactt cattaccagg
     1501 atgtgattgg ggaagagttg aaggaacagc agttggattt tcatttttct cacactcaat
     1561 ttacggaggc ggaggaccag gagtttttaa tggaaatcac gttgttacaa gacactcatc
     1621 aggaatggca attccatgtg ttgcagttgc agttgcatta gatgcaggaa ctcaaatgtt
     1681 ttcaccagaa tcaacatcag caattgtttt agatactttt caggatgtgc caatcatgat
     1741 gaacccactc aaggaggtcg cgaaagcggt gtaaagagga attgtttcat ggttacaact
     1801 acagattcag atccagttca atacgaaatt tttccacaga gaatgttaga tccatcatca
     1861 gcacaagaac ttcttaacag attagaacag attgaaggag ttactagagt tttagttcaa
     1921 ggaccaagat taccattaac agttccatca ggaccaggaa aaggagttcc agttgatcac
     1981 caggcaagag aagttattag agttggagat caagttatgg aacttcaggt taaagttgga
     2041 agaattagat tagaagcaga aaacgatgtt atggaagaag ttagagcact ttgtgaagaa
     2101 ctttttccat ttccatttga acttaaagaa ggattatttc ttagaagaaa agcaactgtt
     2161 acagattacg caaaatacgg accagatgaa atggttgaag ataaagatat ttacggaatg
     2221 actgatccga acatgtcgcc gactgcgaag atctgcgagt catgcgtcga cacgagggag
     2281 taatatgttt ggaagagaaa cacaggttat tgattgtaga gaaggaatgg gattaggaaa
     2341 aggtggagga ttagcacaaa gaggaacatt atcagaaact ggaagaccag atgttattgc
     2401 aattgcaatg tcaccaggaa gaagacacat tacaaaacca gtttgtgaaa ttactcacgg
     2461 attaagaaga gaaggaattc aggtttcagt tttagtttta gaagcaggag caggaattcc
     2521 agatgaagga ccatcagata gaatggcaaa aggatacgga ccaaaatttg gacttacaca
     2581 aaaagaaatt gaacagattg caagacacag agttgcactt attcaccttg gaaacattga
     2641 ttcacacgtt atttacaaaa ctaaatacgt tcttagatac gttgatattc cagcagttat
     2701 tgcatgtcaa aacccagttg attttgaaga ttttgcaaaa gcaggaatta aaactagatt
     2761 agttatgcca atggataaag atattgatac acaaggaact gttatggaaa ttgttgcagg
     2821 aattacaaga ggagttactt gtcagagagg atcacttaac gaactcgtaa aagcaattat
     2881 gaaaacaata ccacaacata cataaaggag atataattat ggattacaaa gatgatgatg
     2941 ataaaggcgc cagctggtca cacccacaat ttgaaaaagg aggtggatca ggaggtggat
     3001 caggaggttc agcatggtca catcctcagt ttgaaaaatc agcaatggca tatacaccac
     3061 aatattaccc aggagcatca cacgttgcag ttaatagaag aaaacacatg tcaggagatg
     3121 ttgaaaaact tagaacagtt tcagatgatg atttagttgc agcattagga cacagagcac
     3181 caggagcaga ttatccatca actcacccac cattagcaga aatgggagaa ccagattgtc
     3241 cagttagaca aatggttgaa ccaacaccag gagcagcagc aggagataga gttagatact
     3301 cacagtttac tgattcaatg tattcagcac catcaattcc atactttaga tcatactacg
     3361 cagcaattaa ctttagagga gttgatccag gaacattatc aggaagacaa attgttgaag
     3421 caagagaaag agatatggaa gcacagtgta aagcagcaat tgaatcagaa atgacatgtc
     3481 cagcattagc aggattaaga ggatgtactg ttcacggaca ctcattaaga ttagcagaag
     3541 atggaatgat gtttgatatg cttgaaagaa ctcacttaga aggtggaaac gttattgaag
     3601 ataaagatca ggttggagtt ccaattgata gaaaagttaa ccttggaaaa ccaatgtcag
     3661 atgcagaagc aaagaaaaga acaactattt acagaacaga tggagttaaa tacagatcag
     3721 aagaagaagt tttagatcac gttcacttag ttcaccacag aagaactatg tatggataca
     3781 gaccagaaac agcagcagag acagcaccag gtgtcggacc tgttacatac cacacggtgt
     3841 aaaaggaggt agatacaaat ggcatacaaa tatccatcag agaaactgtt cgttgaggcg
     3901 ctgaaggcaa agtttgcagg aattgatctt gcagatcaga caactaaata cgttagagca
     3961 ggatacgttc aaaacgcaag aaaaagagaa tttcaggcag caggagaaag agttgcagaa
     4021 caaagaggaa ttcaacagta tgatgttaat gttcaccttg gaggaatgac tttaggacaa
     4081 agacagttag ttccatacaa actttcaaca agaccagata ttgttgaagg agatgattta
     4141 cactacgtta ataatccagc aatgcaacag atgtgggatg atatgaaaag aacaattatt
     4201 gttggaatgg atttagcaca cgaaacatta gaaaaacgac ttggaaaaga agttacacca
     4261 gaatcaattg caggatacat ggaagcagtt aatcacacta tgccaggagc agcaattgtt
     4321 caggaacaca tggttgaaac acacccagga ttagttgatg attgttacgt taaaatgttt
     4381 actggagatg atgaacttgc agatgaaatt gattcacaat acgttattaa cattaacgat
     4441 ttatttgata aagaaggaca gaacgaaaaa attaaagcag caattggaaa aacaacttgg
     4501 caagcaattc acattccaac tattgttgtt agatgttgtg atggaggaaa tacatcaaga
     4561 tggtcagcaa tgcaattagg aatgtcatat attgcagcat acaatatgtg tgcaggagaa
     4621 gcagcagttg cagatttagc atttgcagca aaacacgcag cagcagttca gatggcagaa
     4681 atgttaccag caagaagagc aagatcacca aatgaacctg gaggattatc atttggatat
     4741 tgtgcagata tggttcaaac tatgagagtt aaaccagaag atccagtttg gtacacttta
     4801 gaagttgttg catgtggaac aatgttatat gatcagattt ggttaggatc atacatgtct
     4861 ggaggagttg gatttacaca atatgcaact gcagcataca caaacgatgt tttagatgat
     4921 tttacatact acggatacga ttacgcactt aacaaatacg gagatgatgg aactgcacca
     4981 aatgatttag caacagcaac tgatcttgca acagaagtta ctcttaacgg aatggaatgt
     5041 tacgaagatt acccaacatt attagaagat cactttggag gatcacagag agcaggaatt
     5101 ttagcagcag catcagcatg tacaactgga attgcaactg gaaatgcaca agttgcatta
     5161 tcagcatggt atatgtcaat gtacgttcac aaagaaggat ggggaagatt aggatttttc
     5221 ggatacgatt tacaagatca gtgtggagca acaaatgttt gttcatacca gggagatgaa
     5281 ggatgttgtt tagaacttag aggagcaaac tacccaaatt acgcaatgaa tgttggacac
     5341 caaggagaat atgcaggatt tactggagca gcacacgcag gagcacacga tgcatactgt
     5401 tgtaatccat taattaaagt ttgttttgca gatccatcat tagtttttga tttttcatac
     5461 attagaaaag aatacgcaaa aggagcaatg agaacattta gaccagcagg agaaagatca
     5521 ttagttattc cagcaggagt ttaaactagt agcggccgct gcaggatata gcaaaagtgg
     5581 gacttaagtt cccacttttg ctattttata aaaaacgccc tattcgtaat atactattca
     5641 aaactaaata ttcattgaaa tatttgggta ataattaata ttaatctaat atttagatat
     5701 tgagatataa taataacatt tttttaaaaa atcatctatt ttcatcttaa ttttatatat
     5761 taaagcccat tttttgagta ttcaaattca aattattgtg ttattaacat cttatatata
     5821 aacttttcta tttaatgtta atgaaaaagt gaatatatat acatagagta atgttatgat
     5881 gtatatatca aaaaaaacta gaggagacga ccttccatga cagaatataa accaacagtt
     5941 agattagcaa caagagatga cgtacctagg gcagtaagaa ctcttgcagc agcttttgca
     6001 gattatccag caacaagaca cacagttgat ccagatagac acattgaaag agttacagaa
     6061 ttacaagaat tatttttaac aagagttggt cttgatatag gtaaagtatg ggttgcagat
     6121 gatggagcag cagttgcagt ttggacaaca ccagaatcag ttgaagctgg tgctgttttt
     6181 gcagaaattg gaccaagaat ggctgaatta tcaggatcaa gattagcagc tcagcaacag
     6241 atggaaggat tacttgcacc acacagacca aaagaaccag catggttttt agcaacagtt
     6301 ggagtttcac cagatcacca gggaaaaggt ttaggttctg ctgttgtact tcctggtgtt
     6361 gaagcagctg aaagagcagg agttccagca tttttagaaa catcagcacc aagaaatctt
     6421 ccattttatg aaagacttgg atttacagtt acagcagatg ttgaagttcc agaaggacca
     6481 agaacatggt gtatgacaag aaaaccagga gcataaccat ggctccgacc gaagccaccc
     6541 ggggcggccc cgccgacccc gcacccgccc ccgaggccca ccgcggggga cacaccgaac
     6601 acgccgaccc tgctgaacac gcggcgcagt tcggtgccca ggagcggatc gggaattaat
     6661 tcgaagctgc tggtgaaaga gaccctatct tacctgctaa aatctaagtt aattactaat
     6721 ttattattaa tttattatta gattgggtaa aatagtaaaa gaaaactaaa ggaaacctaa
     6781 tatggtttcc tttttttata tatttttaat tcactggggg gatttaaaac ttcattttta
     6841 atttaaaagg atctaggtga agatccttgg tgcactctca gtacaatctg ctctgatgcc
     6901 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     6961 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     7021 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     7081 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     7141 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     7201 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     7261 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     7321 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     7381 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     7441 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     7501 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     7561 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     7621 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     7681 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     7741 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     7801 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     7861 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     7921 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     7981 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     8041 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     8101 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     8161 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     8221 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     8281 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     8341 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     8401 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     8461 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     8521 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     8581 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     8641 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     8701 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     8761 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     8821 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     8881 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat ggtacccatt
     8941 atttagtttt tttttatcac cacaacgtat aaaccccctt tcagaagaaa acagagaatg
     9001 aaaacagatt ttaatttata tatcgtatct ataatttagt ttttttttat tgcagtttta
     9061 gaagtgcgtt tgtcctggag cgaaatacac gatatcgtgt agtctccttc agatgaaaat
     9121 aatattttta attttcagaa atattactgg aaaaatacct gaaaaatatt tgaaaaaaaa
     9181 acatacaccc agcggggaaa aatcgaacat tttgaaaaag tctatttata gggtttaaac
     9241 agggccaaga cctgcgggat ataccccaaa aatgttcgat aaaaaaatcg aacatttttc
     9301 gaacattttt tataatgttc gatgtgtgcg agtatgttcg ataaaaaaat cgaacatttt
     9361 gaaagacaat ttgaagagat ataaaaaaaa taatctatta taatatagat atataatatg
     9421 ttcgatatgt tcgatat
//
